Reverse transcriptase (RT) is an enzyme required for replication of the Human Immunodeficiency Virus (HIV) and is the target of most of the therapeutic agents approved for clinical use in the treatment of AIDS. The majority of the drugs that have shown promising clinical results are nucleoside analogs (NRTIs). However, the success of the nucleoside analogs is limited by their toxicity and the ability of the HIV virus to develop drug resistant mutations. A second class of drugs that also targets RT is the nonnucleoside inhibitors (NNTRIs). There are many unanswered questions regarding the molecular mechanisms of inhibition, drug resistance, and toxicity. This information is crucial for compounds at various stages of development;in early preclinical evaluation to predict and anticipate potential mechanisms of toxicity and drug resistance as well as later in clinical trials and after FDA approval to understand and explain unanticipated effects in a clinical setting. A knowledge of how these compounds interact with wild type and mutant forms of the viral polymerase (HIV-1 RT) and the cellular polymerase (human mitochondrial DNA polymerase 3 associated with NRTI toxicity) providing insight for mutant forms of RT associated with drug resistance and mutant forms of mtDNA pol 3 implicated in NRTI toxicity are essential for the design of more effective, less toxic therapies. Moreover, detailed molecular studies serve to complement/augment an understanding of drug modes of inhibition and toxicity at a cellular level and in the clinic. The current proposal has two broad objectives. The first will provide key mechanistic information regarding NRTI mode of inhibition and drug resistance for the viral polymerase and potential toxicity for the cellular mitochondrial polymerase. The second objective builds on a new direction in our laboratory to combine our detailed mechanistic studies and cellular/virological assays with collaborative studies using computational modeling and organic synthesis to discover novel inhibitors of RT. We have a productive and cohesive group and our program is set apart by these interlinked disciplines. This multidisciplinary strategy puts us in a unique position for providing detailed mechanistic information at the enzyme and cellular level and valuable input for inhibitor design as well as answering unsolved and important questions regarding drug toxicity, mode of inhibition, and drug resistance. All of these studies will provide information crucial for the design of better therapeutic agents and strategies for the treatment of AIDS. PUBLIC HEALTH RELEVANCE: The World Health Organization estimates that at end of 2007 over 42 million people worldwide are infected and this number is growing. There continues to be a significant need for new drugs and drug combinations to combat this disease. The studies outlined in this proposal will combine mechanistic studies with computational guidance to design more effective HIV therapies that have improved therapeutic properties.